Study On Secondary Metabolites From Endophytic Fungus And Marine Fungi And Their Biological Activities

As an important biological resource producing natural products,fungal resources are increasingly valued by researchers towards the development of lead compounds with medicinal value.In order to screen out the structural molecules with novel structures and significant activity from fungi,this thesis systematically and thoroughly studied the secondary metabolites of an endophytic fungus Trichoderma koningiopsis A729 as well as two marine fungi Phomopsis tersa FS441 and Trichobotrys effuse FS524,which were derived from Morinda officinalis and marine sediment sample,respectively.The fermentate products of the three strains were separated and purified by various chromatography techniques such as silica gel columns,ODS,Sephadex LH-20,and high-performance liquid chromatography.Furthermore,the planar structures and absolute configurations of these compounds were determined by analysis of 1D and2D NMR,HRESIMS,IR,UV,optical rotation,CD,coupling with single crystal diffraction spectroscopic data.A total of forty nine compounds were isolated and identified,including four new types of skeletons,twenty seven new compounds,four pairs of new enantiomers,and two new natural products.Their structural types mainly involved diterpenes,sesquiterpenes,meroterpenoids,norsteroid,pyridone alkaloids,and phenol derivatives.Structurally,among the metabolites isolated from the endophytic fungus T.koningiopsis A729,compounds A1 and A2 were intriguing diterpene alkaloids possessing guanacastane skeleton,the difference was that compound A2 possessed a rare 6/7/5/5/7 pentacyclic alkaloid skeleton,while compound A3 contained a unique highly fused 4/7/5/6 tetracyclic ring system the formation of a spiral skeleton.Among the metabolites of marine fungus P.tersa FS441,compound B1 was the first report of tropolonic sesquiterpene consisting of a highly fused 6/6/11/6/5/5 ring system,compound B2 possessed an unusual 7/6/11/6-5 spiral core skeleton,while compounds B3 and B4 were the first examples of phenolic sesquiterpene meroterpenoids featuring a highly fused 6/6/6/6 tetracyclic ring system.In addition,compound B5 was a novel norsteroid with an unusual 6/5-5 tricyclic skeleton.Compounds B6-B8 and B10 were isolated as four pairs of phenylfuropyridone racemates with the rare naturally occurring 6-6/5/5 ring system,while compound B9 was discovered as phenylpyridone racemate with a 6-6/6 ring system.Furthermore,compound B12 represented the first member of 5-phenylpyridone derivatives with an unprecedented furo[3,2-c]pyridin-4?5H?-one skeleton.Moreover,among the metabolites of marine fungus T.effuse FS524,compounds C1 and C2shared an unique 6-6/6/6 tetracyclic ring system with the formation of an unprecedented ploy-substituted 9-phenyl-hexahydroxanthone skeleton,which was often encountered as one of the most ubiquitous and intriguing functional moieties in the pharmaceutical drugs but rarely discovered in natural products.The cytotoxic activities of compounds A1-A10,B1-B18,B20,B22-B23,and C1-C6 were tested by the SRB method,and the antibacterial activities of compounds A1-A2,A4-A6,A9-A10,and B6-B12 were tested by standard microdilution method.Furthermore,compounds C1-C6 were tested for the inhibitory activity of nitric oxide?NO?production in lipopolysaccharide?LPS?-induced RAW 246.7 mouse macrophages using the Griess method.The results of cytotoxic experiments showed that new compound B2 showed significant antiproliferative effect against four tumor cell lines with an IC₅₀0 value of 0.50-1.30?M,which were much better than that of the positive control adriamycin?1.10-1.50?M?.In addition,compounds B1,B3-B4,and B10b showed moderate inhibitory activities against four tumor cell lines.The results of antibacterial activities revealed new compounds A1 and A2 showed potent antibacterial activities against B.subtilis with MIC values of 10 and 2?g/mL,respectively.Furthermore,compounds B10a and B10b demonstrated specific antibacterial activities against S.aureus.The results of NO inhibitory activities indicated new compounds C3 and C4 showed moderate NO inhibitory activities in RAW264.7 macrophages induced by LPS.Through the study of secondary metabolites from an endophytic fungus and two marine fungi,three types of highly fused new skeleton meroterpenoids,one type of highly oxygenated new skeleton norsteroid,together with a series of polycyclic system diterpenoid alkaloids,pyridone alkaloids and highly substituted phenol derivatives were isolated and determined.Molecules with significant activity against B.subtilis and potent tumor cytotoxicity were screened out.This paper not only enriched the structural diversity of secondary metabolites from endophytic fungi and marine fungi,but also provided important model structures and prodrugs for the research and development of new drugs,laying an important material foundation for the future development of innovative drugs with China's independent intellectual property.